This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The HIV-1 Gag protein directs the highly orchestrated process of particle assembly and budding. During assembly, the uncleaved Gag polyprotein interacts with the viral RNA and the envelope glycoprotein complex (Env) to coordinate the production of infectious virions. The cellular trafficking machinery is recruited by HIV-1 Gag and participates in essential steps of particle assembly and budding. We identified a novel interaction between Gag and adaptor protein complex AP-3. This interaction directs Gag trafficking to multivesicular bodies (MVB). Disruption of Gag-AP-3 interaction prevents Gag from reaching to MVBs and inhibits particle formation. We have identified several Gag-binding proteins by screening human cDNA library. Recently, we are investigating their roles in HIV assembly using variety of approaches such as lentiviral mediated gene transfer, RNAi-mediated gene knockdown and site-directed mutagenesis. These studies not only have broader biological implications in understanding the association between cellular factors and viral proteins during HIV assembly, but also provide us new information to design drugs to combat the virus.